Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Concept Videos

Protein-Drug Binding: Determination Methods01:22

Protein-Drug Binding: Determination Methods

648
Determining protein-drug binding can be achieved through indirect and direct methods, each providing valuable insights into the interaction between proteins and drugs.
Indirect methods involve isolating the bound drug from its free form in biological samples such as blood, serum, or plasma. These techniques aim to measure the percentage of drugs bound to proteins. Equilibrium dialysis is a commonly used method where the free drug concentration at equilibrium is measured by separating the bound...
648
Affinity and Avidity01:41

Affinity and Avidity

38.7K
Overview
38.7K
The Equilibrium Binding Constant and Binding Strength02:18

The Equilibrium Binding Constant and Binding Strength

15.0K
The equilibrium binding constant (Kb) quantifies the strength of a protein-ligand interaction. Kb can be calculated as follows when the reaction is at equilibrium:
15.0K
Electron Affinity03:07

Electron Affinity

43.3K
The electron affinity (EA) is the energy change for adding an electron to a gaseous atom to form an anion (negative ion).
43.3K
Factors Affecting Protein-Drug Binding: Protein-Related Factors01:20

Factors Affecting Protein-Drug Binding: Protein-Related Factors

551
Drug binding to proteins is a key aspect of pharmacokinetics and can influence a drug's distribution, absorption, and elimination in the body. Several factors, including the drug's physiochemical properties, protein concentration, disease states, and the number of binding sites on the protein, influence this process.
The physicochemical properties of a drug play a significant role in its ability to bind to proteins. Lipophilic drugs, which dissolve in fats, oils, and lipids, can be...
551
Conjugated Proteins02:50

Conjugated Proteins

28.6K
Simple proteins and protein complexes contain only amino acids. In contrast, many other proteins, called conjugated proteins, covalently bond with non-protein moieties.
Nucleoproteins are protein complexes that contain nucleic acids, categorized as deoxyribonucleoproteins (DNPs) or ribonucleoproteins (RNPs) respectively. The nucleosome is a typical example of a DNP where nuclear DNA is associated with histone proteins. The major antigen for the Covid-19 virus SARS-CoV is an RNP that is critical...
28.6K

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Correction to "Monooxygenase Activity of Indoleamine 2,3-Dioxygenase".

Journal of the American Chemical Society·2026
Same author

Label-free optical observation of disordered-to-ordered transitions in single intrinsically disordered proteins.

Npj biosensing·2026
Same author

Monooxygenase Activity of Indoleamine 2,3-Dioxygenase.

Journal of the American Chemical Society·2026
Same author

Split-APEX implicates splicing factor SRSF1 and splicing helicases in ribosomal biogenesis.

Frontiers in molecular biosciences·2026
Same author

Tracking single-molecule ferritin reassembly and disassembly using polymer-coated nanopores.

Nanoscale·2025
Same author

Multi-photon fluorescence-lifetime imaging of a genetically-encoded heme sensor.

Journal of inorganic biochemistry·2025

Related Experiment Video

Updated: Jan 28, 2026

Protein Purification-free Method of Binding Affinity Determination by Microscale Thermophoresis
10:22

Protein Purification-free Method of Binding Affinity Determination by Microscale Thermophoresis

Published on: August 15, 2013

31.3K

Precise determination of heme binding affinity in proteins.

Galvin C-H Leung1, Simon S-P Fung1, Nicholas R B Dovey1

  • 1Department of Chemistry and the Leicester Institute of Structural & Chemical Biology, University of Leicester, Leicester, LE1 7RH, United Kingdom.

Analytical Biochemistry
|February 27, 2019
PubMed
Summary

Cellular heme acts as a signaling molecule. A new multivariate curve-resolution (MCR) method precisely determines heme-binding affinity (Kd) by separating component spectra, improving upon traditional spectrophotometric titrations.

More Related Videos

Determining Binding Affinity KD of Radiolabeled Antibodies to Immobilized Antigens
07:39

Determining Binding Affinity KD of Radiolabeled Antibodies to Immobilized Antigens

Published on: June 23, 2022

7.2K
Determining the Ice-binding Planes of Antifreeze Proteins by Fluorescence-based Ice Plane Affinity
08:46

Determining the Ice-binding Planes of Antifreeze Proteins by Fluorescence-based Ice Plane Affinity

Published on: January 15, 2014

9.6K

Related Experiment Videos

Last Updated: Jan 28, 2026

Protein Purification-free Method of Binding Affinity Determination by Microscale Thermophoresis
10:22

Protein Purification-free Method of Binding Affinity Determination by Microscale Thermophoresis

Published on: August 15, 2013

31.3K
Determining Binding Affinity KD of Radiolabeled Antibodies to Immobilized Antigens
07:39

Determining Binding Affinity KD of Radiolabeled Antibodies to Immobilized Antigens

Published on: June 23, 2022

7.2K
Determining the Ice-binding Planes of Antifreeze Proteins by Fluorescence-based Ice Plane Affinity
08:46

Determining the Ice-binding Planes of Antifreeze Proteins by Fluorescence-based Ice Plane Affinity

Published on: January 15, 2014

9.6K

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Spectroscopy

Background:

  • Emerging evidence indicates cellular heme functions as a signaling molecule.
  • Heme interactions with proteins are often transient, differing from classical hemoproteins.
  • Accurate determination of heme-binding affinity (dissociation constant, Kd) is crucial for studying this role.

Purpose of the Study:

  • To develop a precise method for determining heme-binding affinity (Kd).
  • To overcome limitations in spectrophotometric titrations that hinder accurate Kd determination.
  • To characterize holo-protein spectra for previously unstudied heme-binding proteins.

Main Methods:

  • A novel approach using multivariate curve-resolution (MCR) algorithm was employed.
  • MCR separates absorbance contributions from apo-protein, holo-protein, and hemin.
  • Pure component spectra and concentration profiles are estimated for simultaneous fitting to a binding model.

Main Results:

  • The MCR method allows for more precise Kd determination by analyzing concentration profiles of all components.
  • An absorption spectrum for the holo-protein can be calculated, a unique advantage of this method.
  • This approach enables simultaneous fitting to a theoretical-binding model for improved accuracy.

Conclusions:

  • Multivariate curve-resolution (MCR) offers a significant advancement over existing methods for determining heme-binding affinity.
  • The ability to calculate holo-protein spectra provides valuable insights into heme-protein interactions.
  • This method is particularly attractive for characterizing heme binding in proteins where it has not been previously studied.